Interdependent effects of inorganic phosphate and creatine phosphate on sarcoplasmic reticulum Ca2+ regulation in mechanically skinned rat skeletal muscle
Am. Duke et Ds. Steele, Interdependent effects of inorganic phosphate and creatine phosphate on sarcoplasmic reticulum Ca2+ regulation in mechanically skinned rat skeletal muscle, J PHYSL LON, 531(3), 2001, pp. 729-742
1. The effects of creatine phosphate (CP) and inorganic phosphate (P-i) on
sarcoplasmic reticulum (SR) Ca2+ regulation were investigated in mechanical
ly skinned muscle fibres from rat extensor digitorum longus (EDL) muscles.
Changes in [Ca2+] were detected using fura-2 fluorescence, during continuou
s perfusion or when the solution surrounding the preparation was restricted
to approximately 6 mul by stopping perfusion.
2. In solutions with 5 mM ATP and 10 mM CP, stopping the flow for 2-3 min h
ad no effect on [Ca2+] within the bath. This suggests that SR Ca2+ uptake i
s balanced by an efflux under these conditions.
3. In solutions with CP, the introduction of P-i induced a small transient
rise in [Ca2+], due to Ca2+ loss from the SR. Following equilibration with
solutions containing P-i (greater than or equal to5 mM), a maintained decre
ase in [Ca2+] occurred when the flow was stopped. This is consistent with c
alcium phosphate (Ca-P-i) precipitation within the SR, resulting in maintai
ned Ca2+ uptake.
4. In the absence of CP, the [Ca2+] within the bath increased progressively
when the flow was stopped. This rise in [Ca2+] was inhibited by an alterna
tive ATP regenerating system comprising phosphoendolpyruvate (PEP) and pyru
vate kinase (PR). Therefore, the loss of Ca2+ from the SR may result from l
ocal ADP accumulation and the consequent reversal of the SR Ca2+ pump.
5. In the absence of CP, the initial Ca2+ release associated with the intro
duction of P-i increased markedly. Following prolonged equilibration with s
olutions containing P-i, a rise in [Ca2+] occurred within the bath when the
flow was stopped. Maintained Ca2+ uptake associated with Ca-P-i precipitat
ion was not apparent at any level of P-i tested (1-60 mM), when CP was abse
nt.
6. These results suggest that withdrawal of CP is associated with activatio
n of a SR Ca2+ efflux pathway. This may involve reversal of the SR Ca2+ pum
p, due to local ADP accumulation. In the absence of CP, the dominant influe
nce of P-i appears to involve further Ca2+ efflux via the SR Ca2+ pump. The
possible relevance of these effects to skeletal muscle fatigue is consider
ed.